Death after intrathecal capsaicin?

Experimental Techniques
1

Hi there,

I’ve been conducting lately intrathecal capsaicin injections to ablate TRPV1 fibers. I inject 7’5ul of a 2ug/ul capsaicin solution in 0.9% saline/10%ethanol/10%Tween80 in adult C57 mice (7weeks old). Some of them die right after injection: the breathing turns irregular and then die. At first I thought it was a volume problem, that I was causing to much pressure in the CSF, but animals injected with vehicle at same volume/speed/etc never die. All mice are anesthesized with isofluorane during procedure and always check site of injection with tail-flick.

Anyone with a cue of what can I be doing wrong? I’ve searched the internet but I cannot find anyone reporting this.

Thanks!

2

Hi Patricia,
Thanks for bringing this question to the group. A question - what spinal level are you injecting at?
Also, have you done just vehicle injections? Do you get the same outcome?

3

Hi Alex,

I’m injecting through L5-L6 vertebral space with a 25ul Hamilton syringe with 30g needle. I injected 18 animals with the capsaicin solution and 5 died, whereas 12 out of 12 injected with just vehicle lived, so I assume it has something to do with the capsaicin and not with the injection procedure… But I’m clueless about how capsaicin could provoke death.

Thanks for your response!

4

OK well then yes, it’s likely the capsaicin. Maybe there is some massive sympathetic response that causes a cardiac death? I’m not sure. I’m sure someone has seen this before but it’s the kind of thing that doesn’t make it into the paper. @thicunha @tberta @tonellor?

5

The dose of capsaicin seems to be 15 micro/site, which is quite high. I agree with Alex that a sympathetic response may be the cause of death. When we were using Resineferatoxin, we saw a robust sympathetic activation after systemin injection which sometimes led to death. Please see our paper:

Hyperactivation of sympathetic nerves drives depletion of melanocyte stem cells.
Zhang B, Ma S, Rachmin I, He M, Baral P, Choi S, Gonçalves WA, Shwartz Y, Fast EM, Su Y, Zon LI, Regev A, Buenrostro JD, Cunha TM, Chiu IM, Fisher DE, Hsu YC. Nature. 2020 Jan;577(7792):676-681.

6

Hi Thiago,

I inject only 7’5ul per animal, which is a quantity that is in the range reported by papers that use intrathecal capsaicin at that concentrations. I though that dose was safe but now knowing that systemic capsaicin analogs can lead to death because of sympathetic activation it is reasonable to think that something like that happened to my animals. Maybe some of them are specially sensible for some reason… I will try with smaller quantities to see if the survival rate is better.

Thank you for your insights, I definitelly will check the paper.
And thank you Alex for your help.
If anyone in the community has any more info/recommendation it will be very much appreciated.

Thanks!

7

Very interesting to hear this, I haven’t heard of this happening in adult animals before. Is it possible the capsaicin is getting out of the spinal column? In the studies using systemic capsaicin injection in neonates to kill off the C-fibers (before TRPV1 was identified), it was common to see >50% mortality-the pups would turn blue and die. Sounds like it could be an interesting paper to characterize the sensory-sympathetic loop if anyone is willing to do the work. This has been reported in the bone innervation field, but not well characterized. There is also supposed to be a peptidergic sensory projection to the spinal intermediolateral cell column (preganglionic sympathetics), as well as a brainstem loop, and reports that vascular TRPV1+ innervation regulates vascular tone. Not sure how to get around it though. I would email Alan Basbaum and ask him.